Diabetes mellitus (DM) is characterized by a perturbed hormonal milieu, including hypergluconemia, that eventually leads to metabolic deterioration. Although glucose and amino acids (AA) can directly affect glucagon (G) secretion, progress toward understanding the molecular mechanisms underlying nutritional-regulation of G gene expression has been hampered by the absence of a homogeneous a-cell population. The recent creation of the alphaTC9 cell line circumvents the confounding effects presented with previous methodologies. Our initial research objectives are to characterize nutritional and hormonal regulators of G gene transcription and G secretion in alphaTC9 cells. Glucose (O, 2.5, and 16.5 mM) and 11 individual AA (O, 5, and 10 mM) will be tested in the presence or absence of various hormonal effectors of G secretion. Combinations of the AA will be similarly tested. To identify nutrient-control of G mRNA expression, standard molecular tests including Northern analyses, promoter activity assays, and analyses of DNA-protein interactions will be performed. Information gained from this project will be essential for identifying and characterizing specific DNA-binding regulatory proteins, and for developing non-metabolizable nutritional analogs to abate the hypergluconemia associated with DM pathophysiology.